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Direct Supercritical Angle Localization Microscopy for Nanometer 3D Superresolution

View ORCID ProfileAnindita Dasgupta, View ORCID ProfileJoran Deschamps, View ORCID ProfileUlf Matti, Uwe Hübner, Jan Becker, View ORCID ProfileSebastian Strauss, View ORCID ProfileRalf Jungmann, View ORCID ProfileRainer Heintzmann, View ORCID ProfileJonas Ries
doi: https://doi.org/10.1101/2020.06.25.171058
Anindita Dasgupta
1Cell Biology and Biophysics, European Molecular Biology Laboratory, Heidelberg, Germany
2Leibniz Institute of Photonic Technology, Jena, Germany
3Institute of Applied Optics and Biophysics, Friedrich-Schiller-University, Jena, Germany
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Joran Deschamps
1Cell Biology and Biophysics, European Molecular Biology Laboratory, Heidelberg, Germany
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Ulf Matti
1Cell Biology and Biophysics, European Molecular Biology Laboratory, Heidelberg, Germany
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Uwe Hübner
2Leibniz Institute of Photonic Technology, Jena, Germany
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Jan Becker
2Leibniz Institute of Photonic Technology, Jena, Germany
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Sebastian Strauss
5Faculty of Physics and Center for Nanoscience, Ludwig Maximilian University, Munich, Germany
6Max Planck Institute of Biochemistry, Martinsried, Germany
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Ralf Jungmann
5Faculty of Physics and Center for Nanoscience, Ludwig Maximilian University, Munich, Germany
6Max Planck Institute of Biochemistry, Martinsried, Germany
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Rainer Heintzmann
2Leibniz Institute of Photonic Technology, Jena, Germany
4Institute of Physical Chemistry and Abbe Center of Photonics, Friedrich-Schiller-University, Jena, Germany
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Jonas Ries
1Cell Biology and Biophysics, European Molecular Biology Laboratory, Heidelberg, Germany
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  • For correspondence: jonas.ries@embl.de
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Abstract

3D single molecule localization microscopy (SMLM) is an emerging superresolution method for structural cell biology, as it allows probing precise positions of proteins in cellular structures. Supercritical angle fluorescence strongly depends on the z-position of the fluorophore and can be used for z localization in a method called supercritical angle localization microscopy (SALM). Here, we realize the full potential of SALM by directly splitting supercritical and undercritical emission, using an ultra-high NA objective, and applying new fitting routines to extract precise intensities of single emitters, resulting in a four-fold improved z-resolution compared to the state of the art. We demonstrate nanometer isotropic localization precision on DNA origami structures, and on clathrin coated vesicles and microtubules in cells, illustrating the potential of SALM for cell biology.

Competing Interest Statement

The authors have declared no competing interest.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-ND 4.0 International license.
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Posted June 26, 2020.
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Direct Supercritical Angle Localization Microscopy for Nanometer 3D Superresolution
Anindita Dasgupta, Joran Deschamps, Ulf Matti, Uwe Hübner, Jan Becker, Sebastian Strauss, Ralf Jungmann, Rainer Heintzmann, Jonas Ries
bioRxiv 2020.06.25.171058; doi: https://doi.org/10.1101/2020.06.25.171058
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Direct Supercritical Angle Localization Microscopy for Nanometer 3D Superresolution
Anindita Dasgupta, Joran Deschamps, Ulf Matti, Uwe Hübner, Jan Becker, Sebastian Strauss, Ralf Jungmann, Rainer Heintzmann, Jonas Ries
bioRxiv 2020.06.25.171058; doi: https://doi.org/10.1101/2020.06.25.171058

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